The focus of this grant is to study the function and regulation of H60. H60 is an NKG2D ligand, and as such, participates in the recognition of virally infected cells and cancer cells by the immunoreceptor NKG2D. NKG2D ligands are expressed on the surface of infected cells to alert the immune system to viral infection, but interestingly are also found on cancer cells but not normal cells. However, the signals that place the NKG2D ligand H60 on the surface of cancer cells are unknown. It has been shown that the interferons down-regulate H60 but not other NKG2D ligands, This proposal centers on the following questions: Do the NF-kB and STAT1 pathways provide opposing activity in the regulation H60 expression? How is the H60 locus involved in the regulation of H60 expression on tumor cells? Does the expression of H60 on tumor cells diminish the efficacy of interferon immunotherapy? To address these questions, the regulation of H60 will be studied in a panel of tumor cell lines with naturally high and low levels of H60 using genetic and pharmacologic reagents that activate and inhibit the NF-kB or STAT1 pathways. In parallel to these experiments, the locus of H60 will be examined for regulatory sites that mediate the cis activation of H60. These experiments are facilitated by preliminary studies which map the genomic locus of H60 from a bacterial artificial chromosome. To examine whether the down-regulation of H60 by interferon limits its anti-tumor effect, cell lines with high and low levels of H60 will be transplanted into mice treated with interferon-alpha. The interferon treatment is expected to inhibit tumor surveillance by natural killer cells only in cell lines that express H60. The goal of these experiments is to generate pre-clinical data to support the use of H60 or related NKG2D ligands as biomarkers to predict the efficacy of interferon immunotherapy. In sum, many cancer cells are recognized by the immune system because they display NKG2D ligands at the cell surface which identify the cancer cell as an abnormal cell. This grant proposes to understand the function and regulation of the NKG2D ligand H60. An understanding of how H60 is placed on the tumor cell surface can help to enhance the immune-mediated detection and destruction of cancer.